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SEPGuides: Science Education Partnership: Bacterial Transformation Kit

SEPGuides is the go-to place for all SEP kit protocols and activities, as well as the posting area for announcements and program updates. If something is missing or you have an idea to contribute, please e-mail us at sep@fredhutch.org

Painting with Bacteria

Making LB Agar from Powder

For Supplies-Only requests, you have the option to order premade LB agar bottles (like those provided in the kits) or LB agar powder for making it in your classroom using a microwave. Click here for directions on how to make the LB agar from powder.

Frequently Asked Questions

Ampicillin---Storage/Stability

The stability of ampicillin solutions is dependent on temperature and pH. Ampicillin solutions should not be autoclaved. Stock solutions (50 mg/ml) should be sterilized by filtration through a 0.22 μm filter.

Ampicillin solutions can be added to agars or culture media which have been autoclaved and cooled to 45-50 °C. Culture plates with ampicillin can be stored at 2-8 °C for up to two weeks.

Stock solutions may be stored at 2-8 °C for up to 3 weeks. For long term storage (4-6 months), stock solutions should be stored at -20 °C. At 37° C in culture, ampicillin is stable up to 3 days. 

Additional Web Resources

Resources on the wide world of bacteria worth checking out.

Visual Lab Guide

Kit Description

Introduction:

Students are hired as employees of “Genes-R-Us” to work on a procedure for transforming pBLU and/or pGLO plasmid DNA into E. coli. This is great lab for teaching students that DNA codes for instructions that give cells visible characteristics. Teachers have the option to let students work on optimizing the transformation procedure as a second component to the lab.

Explanation of Plasmids:

pBLU is a plasmid that contains the gene for beta-galactosidase (lactase) also referred to as lacZ. In the E.coli we use, JM101, the lac operon has been partially deleted. With the pBLU plasmid, the lactase will work on X-gal (a compound consisting of galactose linked to an indole) cleaving the compound resulting in a insoluble indigo colored compound. This is called the blue-white screen because it acts as a visual indication of a functional lacZ transformation. For pBLU transformation you would need ampillicin/X-gal/LB agar plates.
 
pFLO is plasmid that contains genes derivative of GFP (green fluorescent protein) also referred to as mGFP or modified GFP. After a pFLO transformation, fluorescent proteins are expressed by the cells that can be visualized under white light or glow under UV light. Because the expression of the mGFP does not require lacZ, X-gal is not required. For pFLO transformation you would need ampillcin/LB agar plates

  • SEP has 4 Transformation Kits.
  • Each kit has 2 crates and an incubator.
  • Teacher must have a heat source to melt agar, ethanol or bleach for cleaning, and ice.
  • SEP teachers who have their own equipment may request supplies only. Fill out and email the Supplies Request Form to sep@fredhutch.org.

Bacterial Transformation Landing Page